S-SAD phasing study of death receptor 6 and its solution conformation revealed by SAXS

Abstract

A subset of tumour necrosis factor receptor (TNFR) superfamily members contain death domains in their cytoplasmic tails. Death receptor 6 (DR6) is one such member and can trigger apoptosis upon the binding of a ligand by its cysteinerich domains (CRDs). The crystal structure of the ectodomain (amino acids 1-348) of human death receptor 6 (DR6) encompassing the CRD region was phased using the anomalous signal from S atoms. In order to explore the feasibility of S-SAD phasing at longer wavelengths (beyond 2.5 Å ), a comparative study was performed on data collected at wavelengths of 2.0 and 2.7 Å . In spite of sub-optimal experimental conditions, the 2.7 Å wavelength used for data collection showed potential for S-SAD phasing. The results showed that the R ano /R p.i.m. ratio is a good indicator for monitoring the anomalous data quality when the anomalous signal is relatively strong, while d 00 /sig(d 00 ) calculated by SHELXC is a more sensitive and stable indicator applicable for grading a wider range of anomalous data qualities. The use of the 'parameter-space screening method' for S-SAD phasing resulted in solutions for data sets that failed during manual attempts. SAXS measurements on the ectodomain suggested that a dimer defines the minimal physical unit of an unliganded DR6 molecule in solution.Received 20 January 2012 Accepted 2 February 2012 PDB References: death receptor 6, data set A1, 3u3p; data set A2, 3u3q; data set B1, 3u3s; data set C1, 3u3v; data set D1, 3u3t.

Structure of the Leanyer orthobunyavirus nucleoprotein–RNA complex reveals unique architecture for RNA encapsidation

“…Only LEAV N with RNA of bacterial origin and the reconstituted LEAV N with synthesized ssDNA complex crystallized and diffracted X-rays beyond 3.0 Å. The structure of the binary complex of LEAV N with ssRNA, crystallized in P1 space group, was solved by the Se-single wavelength anomalous dispersion (Se-SAD) method (27) using X 2 DF HT structure determination pipeline (28,29) and refined to 2.78 Å resolution with an R factor of 23.33 (R free = 27.40). The LEAV N-ssDNA complex, crystallized in P1 space group as well, was solved by molecular replacement (30) using the LEAV N-ssRNA structure as a template and refined to 2.68 Å resolution with an R factor of 22.27 (R free = 24.30) (Table S1).…”

“…The LEAV N-ssRNA complex crystallized in space group P1 with unit cell parameters of a = 49.84, b = 73.83, and c = 79.88 Å. The initial phases were determined using X 2 DF structure determination pipeline (28,29) by the Se-SAD method (27) and the initial model was built by PHENIX AutoBuild (41). The LEAV N-ssDNA complex structure was solved by Molecular Replacement (301) using the solved LEAV N-ssRNA structure as a search model.…”

Combination Therapy of LysGH15 and Apigenin as a New Strategy for Treating Pneumonia Caused by Staphylococcus aureus

“…Small-angle X-ray scattering (SAXS) data on full-length LysGH15 were collected at the SIBYLS beamline (Advanced Light Source, Lorenz Berkeley National Lab, USA), as described in a previous report, with some modifications (26). Briefly, each sample was measured with four exposures (0.5, 1, 2, and 4 s) at 10°C at three concentrations (2.5 ϫ 10 3 , 5.0 ϫ 10 3 , and 1.0 ϫ 10 4 g/ml) in a buffer composed of 20 mM Tris-HCl (pH 7.5), 500 mM NaCl, and 5% glycerin.…”

Role of death receptors in the regulation of hepatocyte proliferation and apoptosis during rat liver regeneration

“…This initiates the recruitment of adaptor proteins such as Fas-associated death domain (FADD) or TNF-receptor-associated death domain (TRADD). Recruiting FADD initiates the assembly of the death-inducing signaling complex, thereby facilitating apoptosis (Ru et al, 2012). Recruiting TRADD results in the activation of the nuclear factor kappalight-chain-enhancer of activated B cells (NK-κB), JNK, and p38 MAP kinase pathways, which regulate several essential cellular activities such as proliferation, differentiation, and apoptosis.…”